Takaaki Arigami

Evaluation of sentinel node concept in gastric cancer based on lymph node micrometastasis determined by reverse transcription-polymerase chain reaction.

Objective:To determine the adequacy of sentinel node (SN) concept based on micrometastasis using immunohistochemistry (IHC) and reverse transcription-polymerase chain reaction (RT-PCR) in gastric cancer. Summary Background Data:The SN concept has recently been introduced in gastrointestinal tract cancers. The precise detection of lymph node metastasis including micrometastasis is important for SN navigation surgery. Methods:Sixty-one patients with gastric cancer who were preoperatively diagnosed with T1-T2 (cT1-T2) and N0 (cN0) were enrolled. They underwent standard radical gastrectomy with lymph node dissection. One day before surgery, 4 mCi of 99mTechnetium-tin colloid was endoscopically injected into the submucosa around the tumor. During surgery, radioisotope uptake in the lymph node was measured using Navigator GPS. All dissected lymph nodes were examined by RT-PCR in addition to hematoxylin and eosin staining and IHC. Results:Sentinel nodes were identified in all patients (100%). The incidences of metastasis determined by hematoxylin and eosin and IHC were 8.2% (5 of 61) and 13.1% (8 of 61), respectively. Micrometastases undetectable by IHC were identified in 14 patients (23.0%) by RT-PCR. Only 1 patient had micrometastasis detectable by RT-PCR in lymph nodes other than SN, but this patient had a cT2 tumor. In patients with cT1 and cN0 tumors, the false negative and accuracy rates were 0% and 100%, respectively. Conclusions:Although the incidence of micrometastasis detected by RT-PCR was quite high, SN navigation identified such metastasis in all patients except one. Thus, the SN concept was applicable to patients with cT1 and cN0 gastric cancer, even when micrometastasis was detectable by RT-PCR.

Lymphatic invasion using D2-40 monoclonal antibody and its relationship to lymph node micrometastasis in pN0 gastric cancer

The monoclonal antibody D2-40 is a specific lymphatic endothelial markers and D2-40 staining have been applicable to evaluate lymphatic invasion in various malignant neoplasms. In the present study, we investigated lymph node micrometastasis determined by immunohistochemistry (IHC) and reverse transcription–polymerase chain reaction (RT–PCR) in all dissected lymph nodes obtained from 80 patients with node-negative gastric cancer, and analysed the relationship between micrometastasis and clinicopathological findings including lymphatic invasion of the resected primary tumour using D2-40 immunohistochemical staining. The incidence of micrometastasis determined by IHC and RT–PCR was 11.3% (nine out of 80) and 31.3% (25 out of 80), respectively. Although haematoxylin–eosin (HE) staining revealed lymphatic invasion in 11.3% (nine out of 80) of patients, D2-40 staining uncovered new invasion in 23.8% (19 out of 80) of patients. In the diagnosis of HE and D2-40 staining, the incidence of micrometastasis was significantly higher in patients with lymphatic invasion than in those without lymphatic invasion (P=0.0150 and P<0.0001, respectively). Micrometastasis correlated more closely with D2-40 than with HE staining. We demonstrated a high incidence of micrometastasis and lymphatic invasion and a correlation between them even in pN0 gastric cancer. When planning less invasive treatment, the presence of such occult cancer cells should be considered.

Activation of toll-like receptors 2, 3, and 4 on human melanoma cells induces inflammatory factors

Toll-like receptors (TLR) have been shown to be expressed on various types of cancers; however, their functional activity is not known. We examined TLR profiles of human melanoma cells and showed that TLR2, TLR3, and TLR4 were found to be highly expressed. By PCR array analysis, specific stimulation of TLR2, TLR3, and TLR4 on melanoma cells showed significant activation of the adaptor protein MyD88, as well as downstream signal transduction factors nuclear factor-κB and inflammatory response–related factors. Specific ligand activation of TLR2, TLR3, and TLR4 was shown to induce cell migration. Peripheral blood lymphocytes and melanoma purified RNA was shown to activate TLR3 on melanoma cells. These studies show expression and functional activity of specific TLRs on melanoma cells and as potential therapeutic targets to control tumor progression. [Mol Cancer Ther 2008;7(11):3642–53]

Detection of sentinel nodes and micrometastases using radioisotope navigation and immunohistochemistry in patients with gastric cancer

Patients with early gastric cancer may be treated by minimally invasive surgery. This study investigated the value of sentinel node (SN) navigation surgery, including detection of micrometastases, in patients with clinical (c) T1 and T2 gastric cancer.

Functional RET G691S Polymorphism in Cutaneous Malignant Melanoma

RET proto-oncogene encodes a receptor tyrosine kinase whose ligand is glial cell line-derived neurotrophic factor (GDNF), and its polymorphism at G691S juxtamembrane region (RETp) is a germline polymorphism. Cutaneous melanomas, particularly the desmoplastic subtype, are highly neurotropic; thus we sought to determine the frequency of RETp in cutaneous melanoma and its functional responsiveness to GDNF. RETp was assessed in 71 non-desmoplastic cutaneous melanomas (non-DMs) and 70 desmoplastic melanomas (DMs). Melanoma cell lines with RETp, RET wild type (RETwt), BRAF V600E mutation (BRAFmt) or BRAF wild type (BRAFwt) were assessed for functional activity. RETp frequency was significantly higher in DMs (61%) than in non-DMs (31%, P<0.001). BRAFmt was detected in only 11% of DMs. GDNF stimulation significantly amplified cell proliferation, migration and invasion in RETp, but not in RETwt melanoma cells. GDNF stimulation of RETp cell lines enhanced phosphorylation of extracellular signal-regulated kinase (ERK) and Akt of the RET-RAS-RAF-ERK and RET-phosphatidylinositol 3-kinase (PI3K)-Akt pathways, respectively. GDNF response of RETp cells in signal transduction and other functional studies were not affected by BRAFmt. The study demonstrates that RETp is frequently found in cutaneous melanoma, particularly desmoplastic subtypes, and responds to GDNF inducing events favorable for tumor progression.

B7‐H3 expression in gastric cancer: A novel molecular blood marker for detecting circulating tumor cells

The clinical significance of B7‐H3 expression in gastric cancer remains unclear, although the B7 ligand family plays a critical role in the T cell‐mediated immune response. We therefore investigated B7‐H3 expression as a blood marker of circulating tumor cells and determined correlations with tumor progression in patients with gastric cancer. B7‐H3 expression in gastric cell lines was initially evaluated by immunocytochemistry. Furthermore, we used quantitative RT‐PCR to assess B7‐H3 mRNA expression in four cell lines and in 95 blood specimens from patients with gastric cancer, as well as in 21 samples of peripheral blood lymphocytes from healthy volunteers. B7‐H3 expression in cell lines was identified by immunocytochemistry and quantitative RT‐PCR. Blood specimens from patients with gastric cancer contained significantly more copies of B7‐H3 mRNA than those from healthy volunteers without cancer (P < 0.0001). Levels of B7‐H3 expression significantly correlated with overall stage (P = 0.013). The 5‐year survival rate was significantly lower in patients with high B7‐H3 expression than with low expression (P = 0.02). Multivariate analysis demonstrated that B7‐H3 expression was an independent prognostic factor (P = 0.046). Our results indicate that B7‐H3 appears to be a useful blood marker for predicting tumor progression in gastric cancer. (Cancer Sci 2011; 102: 1019–1024)

B7-h3 ligand expression by primary breast cancer and associated with regional nodal metastasis.

Objective: B7 ligand family members have been shown to be important immunoregulatory factors in host tumor immune responses. We hypothesized that B7–H3, a coinhibitory factor, is expressed by primary breast cancer cells and associated with metastasis to regional tumor-draining lymph nodes. Experimental Design: American Joint Committee on Cancer stage I to III primary breast cancers (n = 82) and normal breast specimens (n = 17) were assessed for B7–H3 expression using paraffin-embedded archival tissues. B7–H3 expression by breast cancer cells was assessed by a quantitative real-time reverse transcription-polymerase chain reaction, and B7–H3 protein expression was evaluated using immunohistochemistry. Results: B7–H3 mRNA expression was detected in 32 of 82 (39%) primary breast tumors but not in normal breast tissues (P = 0.0029). B7–H3 expression in primary tumors significantly correlated with increasing tumor size, American Joint Committee on Cancer stage, and lymphovascular invasion (P < 0.0001, P < 0.0001, P = 0.0071). B7–H3 expression was highly correlated to sentinel lymph node and overall number of lymph nodes with metastasis P = 0.003, and P = 0.004, respectively). In a multivariate analysis, B7–H3 mRNA expression of the primary tumor significantly predicted metastasis to regional lymph nodes (P = 0.021, and P = 0.023, respectively). Antibody staining analysis of paraffin-embedded archival tissue breast tumors and flow cytometry of breast cancer cell lines demonstrated B7–H3 protein expression. Conclusions: B7–H3 protein expressed by primary breast cancer cells is a tumor progression factor and is associated with extent of regional nodal metastasis.

Clinical significance of circulating tumor cells in peripheral blood from patients with gastric cancer

The authors hypothesized that circulating tumor cells (CTCs) in patients with gastric cancer are associated with prognosis and disease recurrence. In this study, they evaluated CTCs in gastric cancer and clarified the clinical impact of CTCs.

Clinical Significance of Lymph Node Micrometastasis in Gastric Cancer

Recently, the existence of lymph node micrometastasis (LNM), including isolated tumor cells, has been focused on during the development of molecular diagnostic tools for lymph node metastasis in various malignant neoplasms. In particular, immunohistochemistry and reverse transcription-polymerase chain reaction have been reported to be available for the detection of LNM in gastric cancer. However, at present, the clinical significance of LNM remains unclear in patients with gastric cancer. Therefore, we cannot strategically make light of this issue in clinical management. Currently, minimally invasive treatments, such as endoscopic submucosal dissection and laparoscopic surgery with personalized lymphadenectomy, are widely performed in consideration of postsurgical quality of life (QOL). However, it is important to maintain the balance between QOL and curability when selecting surgical treatments for patients with gastric cancer. If minimally invasive surgery based on LNM status was established for patients with early gastric cancer, it could be performed safely. We reviewed the clinical significance of LNM as an important strategic target in patients with gastric cancer.

Clinical Implication of CXCL12 Expression in Gastric Cancer

Recent research has revealed that tumor cells expressing chemokine receptors have a crucial impact on patient survival. However, there is no information regarding chemokine expression in gastro-intestinal cancer. This study immunohistochemically investigated CXCL12 expression in gastric cancer and evaluated its association with clinical factors, including patient prognosis. A total of 185 gastric cancer patients receiving curative gastrectomy were assessed. CXCL12 expression was evaluated by immunohistochemical analysis. Tumors with CXCL12-positive cancer cells were regarded as CXCL12 positive, and according to the degree of CXCL12 expression, patients were divided into three groups (weak, 31 cases; moderate, 27 cases; strong, 20 cases). Correlations between CXCL12 expression and clinical factors in gastric cancer were then determined. CXCL12 was found in the cellular membrane of cancer cells. Seventy-four of 185 patients were classified into the CXCL12-positive group. Patients were divided into three groups according to the positivity of CXCL12 expression. Significant associations between CXCL12 and lymph node metastases (p < 0.05), depth of invasion (p < 0.01), lymphatic invasion (p < 0.01), tumor diameter (p < 0.05), and clinical stage (p < 0.01) were seen. Univariate analysis revealed that the CXCL12-positive group had significantly poorer surgical outcome than the CXCL12-negative group (p < 0.01). Multivariate analysis revealed CXCL12 to be an independent prognostic factor in gastric cancer (p = 0.02). Cancerous CXCL12 positivity was determined to be an independent prognostic factor in gastric cancer, with CXCL12-positive gastric cancer showing more-aggressive behavior. Autocrine CXCL12 secretion from tumor cells may activate CXCR-4 on the tumor cells, which may be related to of the viability of distant metastases.